The work proposed is based on the Project Leader's previous studies in connection with this Program Project and preliminary data that have been obtained on the role of vasoactive, endothelium-derived mediators in the regulation of microvascular function. We plan to continue the examination of the specific roles of endothelium-derived nitric oxide, prostaglandins and oxygen radical species in vascular reactivity, as well as in shear stress and pressure-induced changes in microvascular tone, in vivo and in vitro. In the context of Specific Aim 1 we will test the hypothesis that the signal transduction pathway that is involved in the transmission of shear forces through microvascular endothelial cells to result in the release of nitric oxide and prostaglandins is related to tyrosine kinase activity, cytoskeletal structures and potassium channels in endothelial cells. We also plan to study the time course of the induction of ecNOS activity in microvessels in response to increases in shear stress. In Specific Aim 2 we will continue our studies of the effects of exercise training on microvascular endothelial cell function in rats. We will test the hypothesis that intermittent increases in blood flow (shear stress) during episodes of exercise activity will result in the upregulation of ecNOS, COX-1 and SOD, and that the increased availability of nitric oxide will affect skeletal muscle oxygen metabolism, all of which will tend to promote increases in blood flow in and work efficiency of exercise-trained skeletal muscle. In Specific Aim 3 the endothelial contribution to changes in arteriolar function due to heart failure will be investigated in coronary and skeletal muscle vessels of dogs and coronary microvessels of humans. We will test the hypothesis that heart failure is associated with a severe reduction in the endothelial synthesis/and or activity in microvessels of both nitric oxide and dilator prostaglandins and that this defect contributes to circulatory collapse. In all three aims responses to agonists, flow and pressure will be studied in isolated, perfused arterioles under controlled conditions and in some instances in vivo, by established methods, including diameter measurements by television microscopy. Nitric oxide will be assayed in vessel perfusates as NO(2), and in plasma as NO(3); cNOS, COX-1 and SOD gene expression and enzyme levels will be quantified by molecular biology techniques and immunohistochemistry.